Enterotoxigenic Escherichia coli (ETEC) strains are categorized as priority B pathogens by NIAID. ETEC-associated diarrhea is responsible for nearly one million deaths each year around the world in addition to causing of illness to immunocompromised individuals and travelers including deployed military personnel. The key virulence factors in ETEC- associated diarrhea are believed to be bacterial adhesins or colonization factors (CFAs) and enterotoxins. Enteroaggregative Escherichia coli heat stable enterotoxin 1 (EAST1) is among the enterotoxins produced by E. coli strains that cause diarrhea in humans and animals. However, the virulence significance of EAST1 in diarrhea has not been defined, in part because a suitable model for key tests has not been identified. Results from our initial studies demonstrated that an additive model using K88ac-receptor positive gnotobiotic piglets and K88ac-fimbrial E. coli model can be readily employed to assess virulence significance of enterotoxins in E. coli-associated diarrhea. In this proposal, we will construct model pathogens expressing EAST1 toxin and K88ac fimbria to define virulence significance of EAST1 in diarrhea and its synergetic contribution to diarrhea caused by E. coli strains that also express other toxins by using the additive piglet challenge model. Determination of EAST1 virulence will provide information for the development of prevention and treatment strategies to protect humans and animals from diarrhea disease. This R15 proposal will recruit promising undergraduate students to be trained with the expert research team as they focus on the biological relevance of EAST1 toxin in diarrhea. [unreadable] [unreadable] Enterotoxigenic Escherichia coli (ETEC) are the primary cause of diarrheal disease. ETEC diarrhea is responsible for 400,000 to 800,000 deaths each year, and brings health concerns and economic consequences in international travelers and military personnel. Enteroaggregative Escherichia coli heat stable toxin 1 (EAST1) is associated with diarrhea. But whether EAST1 toxin is the virulence determinant in diarrhea is unclear. Determination of virulence significance of EAST1 is essential for us to identify other unknown virulence factors in diarrhea. This proposed research is to examine the virulence significance EAST1 in diarrhea. The determination of virulence significance of EAST1 toxin in diarrheal disease will provide essential information to prevent or control this disease. [unreadable] [unreadable] [unreadable] [unreadable]